MCB 2210 001 4 8 2015 Intermediate Filaments and Septins The cytoskeleton consists of at least 4 polymer systems Intermediate Filaments IFs strong rope like polymers of fibrous proteins that resist stretch o 10nm in diameter intermediate between actin and myosin filaments o Very stable disassembly in vitro requires treatment with agents that denature proteins o Do NOT bind nucleotides o Non polar in structure o Can withstand stretching o Play a structural or tension bearing roles in the cell and are important for cell strength and cell cell attachments o Structure Central helical coiled coil rod domain Highly conserved Heads and tails variable in length and sequence Form parallel dimer that assembles into antiparallel tetramer Tetramers assemble end to end to form protofilaments associate laterally to form protofibrils o Futher assemble into an IF Tetrameric subunit has no polarity so the filament has no polarity o Assembly Soluble tetramers can be isolated from cells Suggesting that this is the basic building block of the filament Soluble pool of IF tetramer in cell is very small Slow exchange of IF subunits with the insoluble filaments of the cell The dynamics of these filaments are very modest compared to actin filaments and MTs IF incorporation assay keratins incorporate throughout the filaments MT incorporation assay tubulin subunits incorporate at growing ends o Types of IFs Type I acidic and Type II basic keratins Form heteropolymers composed of acidic and basic subunits 20 keratins in epithelial cells o Keratins are located in epithelial tissue o Different types in different types of epithelia o Origins of cancer can be determined based on the set of keratins found in tumor cells o 10 hard keratins in hair and nails o Keratins in outermost layer of skin become crosslinked together protective layer Type III Vimentin and Vimentin related filaments Form homopolymers that consist of a single IF protein species Vimentin the most widely distributed IF protein o Found in fibroblast endothelial cells white blood cells and embryonic cells Desmin found mainly in muscle cells where it linkes adjacent Z disks of myofibrils Glial Fibrillary Acidic Protein GFAP vimentin related protein found in the glial cells that surround neurons and astrocytes Type IV Neurofilaments Composed of heterpolymers of NF L NF M and NF H Found in axons Important for providing strength in the axon and determining its diameter Nuclear Lamins Nuclear lamina meshwork of IFs that lines the inner surface of the nuclear membrane Composed of lamins A B and C Differ from other IF proteins in that they o Have a longer central rod domain o Contain a nuclear import signal o Assemble into a 2D sheet like lattice o More dynamic than other IFs Disassembly of the nuclear lamina during mitosis is mediated by phosphorylation o Organization and Function of IFs IF binding proteins IFBPs cross link into networks or bundles Network of IFs in the cell is often coincident with the MT cytoskeleton In some cell types disruption of the MT cytoskeleton with MT de stabilizing drugs causes the IF network to collapse Nuclear positioning and support of the nuclear envelope IFs such as vimentin typically extend form the nuclear envelope to the plasma membrane perhaps helping the position of the nucleus Network of lamins at the inner surface of the nuclear membrane helps maintain the structure of the nucleus and is important for nuclear assembly and disassembly Cell cell attachments desmosomes and hemi desmosomes Cells junctions hold together epithelial cells Desmosomes function in cell cell interactions Hemi desmosomes important for the interaction of cells with the extracellular matrix IFs run between desmosomes hemi desmosomes and the nucleus Together IFs and desmosomes provide each cell and the entire epithelium with strength and rigidity Muscle cell integrity Desmin links the Z disks of skeletal muscle together within the myofibril o Connect Z disks to cell junctions in cardiac muscle o Disease Genes encoding IFs are associated with more than 40 clinical disorders Genetic disease epidermolysis bullosa simplex EBS is caused by mutations in keratin genes expressed in skin Skin very sensitive to injury More than 50 mutations in type A lamins Emerey Dreifuss Muscular Dystrophy EDMD Cardiomyopathy Progeria premature aging Septins o 10nm in diameter o Bind and hydrolyze GTP o Found associated with actin filaments and membranes o Associated with arrest in various stages of the cell cycle Cells with mutations cdc in septin genes all arrest at cytokinesis o Structural components of filaments observed at the neck between mother and daughter cells in yeast bud neck o In mammalian cells colocalize with F actin in stress fibers and with Factin in the contractile ring o Organization Individual septin proteins have a GTP binding domain near their N terminus and an helical coiled coil region at their C terminus Thought to form non polar heterohexamers or heterooctamers that can further assemble into filaments rings or cage like structures o Functions Cytokinesis found at cleavage furrow and cytokinetic ring Scaffolding recruiting signaling proteins such as kinases and GTPases that regulate cytokinesis and other aspects of cell function Diffusion Barrier promote compartmentalization of cellular domains Membrane Trafficking movement and fusions of vesicles
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